1. Dorsal Raphe Serotonergic Neurons Control Intertemporal Choice under Trade-off 
Sangyu Xu, Gishnu Das, Emily Hueske, Susumu Tonegawa 
Current Biology 
Volume 27, Issue 20, Pages e3 (October 2017)
DOI: /j.cub
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2. Figure 1 Odor-Guided Intertemporal Choice Task
(A) Task structure. The subject initiates the tasks by sampling odors in the center odor port and then makes a choice by poking in the left or right reward port, waits the prescribed amount of time anywhere in the chamber, and receives a water reward on the chosen side.
(B) Each odor cue was used in six different intensities, corresponding to six different reward delay lengths. O1, odor 1 intensity; O2, odor 2 intensity; DL, delay of left reward; DR, delay of right reward. In this example trial, 70 mL/min of odor 1 is mixed with 10 mL/min of odor 2, indicating a delay of 14 s to the left reward and a delay of 2 s to the right reward.
(C) Raw data from a single example subject performing the OGIC task with equal left and right reward sizes (925 trials, 5 sessions). RD0–RD20, trials in which right reward delay was 0–20 s.
(D) Psychometric curves fitted to choice data in SL1 sessions (10 mice, 64 sessions, 10,882 trials).
(E) Psychometric curves fitted to choice data in SL2 sessions (10 mice, 79 sessions, 10,269 trials).
(F) The subjects chose the left option more often during SL2 sessions (paired t test, t9 = 6.090; ∗∗∗p < 0.001).
(G) The constant in the choice model was not changed during SL2 sessions (Wilcoxon signed-rank test; p > 0.05; n.s., not significant).
(H) The coefficient for the left reward delay was not changed during SL2 sessions (Wilcoxon signed-rank test; p > 0.05).
(I) The coefficient for the right reward delay was not changed during SL2 sessions (Wilcoxon signed-rank test; p > 0.05).
(J) The coefficient for the interaction term was significantly reduced during SL2 sessions (Wilcoxon signed-rank test; ∗∗p < 0.01).
Data are represented as mean ± SEM. See also Figure S1 and Table S1.
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3. Figure 2 Optogenetic Manipulations of DR Serotonergic Neurons
(A and F) Schematics for virus injection and optical implant in DR-Arch (A) and DR-ChR2 (F) subjects.
(B and G) Time points for light delivery in behavior for green light (B) and for blue light (G). Center, center poke transistor-transistor logic (TTL); Side, side poke TTL.
(C) Expression of AAV9-EF1α-DIO-eArch3.0-EYFP in the DR nucleus of a Sert-Cre mouse. Red indicates tryptophan hydroxylase; green indicates Arch. Scale bar, 100 μm.
(D) Psychometric curves for proportion of left choice in the DR-Ctrl group subjected to green light (10 mice, 20,473 trials, 132 sessions). Empty symbols and dotted line indicated light-off trials. Filled symbols and solid line indicate light-on trials. RD0–RD8, trials in which right reward delay was 0–8 s.
(E) Psychometric curves for proportion of left choice in the DR-Arch group (10 mice, 20,785 trials, 137 sessions).
(H) Expression of AAVrh8-CBA-DIO-ChR2-EYFP in the DR nucleus of a Sert-Cre mouse (red indicates tryptophan hydroxylase; green indicates ChR2). Scale bar, 100 μm.
(I) Psychometric curves for the proportion of left choice in the DR-Ctrl group subjected to blue light (10 mice, 20,712 trials, 140 sessions).
(J) Psychometric curves for the proportion of left choice in the DR-ChR2 group (10 mice, 20,455 trials, 147 sessions).
(K) Green light reduced the proportion of left choice in the DR-Arch group (post hoc paired t test; ∗∗p < 0.01) but not in the DR-Ctrl group (post hoc paired t test; p > 0.05). n.s., not significant.
(L) Blue light increased the proportion of left choice in the DR-ChR2 group (post hoc paired t test; ∗∗p < 0.01) but not in the DR-Ctrl group (post hoc paired t test; p > 0.05). n.s., not significant.
(M) There was a positive interaction between green light and the delay interaction term in the DR-Arch group (Wilcoxon signed-rank test; ∗p < 0.05), whereas there was a negative interaction between blue light and the delay interaction term in the DR-ChR2 group (Wilcoxon signed-rank test; ∗p < 0.05).
aq, aqueduct. Data are represented as mean ± SEM. See also Figure S2 and Table S2.
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4. Figure 3
Optogenetic Manipulation of DR Serotonergic Projections in the NAc
(A) Schematic for viral injections.
(B) ChR2-expressing serotonergic projections in a coronal section of NAc. Scale bar, 200 μm.
(C) Schematics for Arch (top) and ChR2 (bottom) virus injection and optical implant, respectively.
(D) Psychometric curves fitted to raw data for the DR-NAC-Ctrl group subjected to green light (10 mice, 20,534 trials, 115 sessions), the DR-NAC-Arch group subjected to green light (10 mice, 20,329 trials, 142 sessions), the DR-NAC-Ctrl group subjected to blue light (10 mice, 20,606 trials, 130 sessions), and the DR-NAC-ChR2 group subjected to blue light (10 mice, 20,506 trials, 150 sessions). Empty symbols and dotted line indicate light-off trials. Filled symbols and solid line indicate light-on trials.
(E) Green light reduced the proportion of left choice in the DR-NAC-Arch group (post hoc paired t test; ∗∗p < 0.01) but not in the DR-NAC-Ctrl group (post hoc paired t test; p > 0.05).
(F) Blue light increased the proportion of left choice in the DR-NAC-ChR2 group (post hoc paired t test; ∗∗p < 0.01) but not in the DR-NAC-Ctrl group (post hoc paired t test; p > 0.05).
(G) There was a positive interaction between green light and the delay interaction term in the DR-NAC-Arch group (Wilcoxon signed-rank test; ∗p < 0.05), whereas there was a negative interaction between blue light and the delay interaction term in the DR-NAC-ChR2 group (Wilcoxon signed-rank test; ∗p < 0.05).
Data are represented as mean ± SEM. See also Table S2.
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5. Figure 4 Trade-off Analysis of Manipulation Effects
(A–D) Heatmaps of predicted light manipulation effects (predicted differences between the proportion of left choice during light-on conditions and the proportion of left choice during light-off conditions) for DR-Arch (A), DR-ChR2 (B), DR-NAC-Arch (C), and DR-NAC-ChR2 (D) subjects. White solid line indicates the perceptual boundary where odor 1 is at the same intensity as odor 2. Red dotted line indicates the valleys and peaks of the predicted manipulation effects. Color bar indicates color coding for levels of effect (unit: proportion of choice).
(E–H) Light manipulation effects plotted against choice difficulty (the distance between average proportion of left choice and either 0 or 1, whichever is closest) and perceptual difficulty (difference between the intensity of the two odor cues normalized by total odor intensity possible) for DR-Arch (E), DR-ChR2 (F), DR-NAC-Arch (G), and DR-NAC-ChR2 (H) subjects and their respective controls.
(I–L) Multiple regression coefficients for choice difficulty and perceptual difficulty accounting for the light effect for DR-Arch (I), DR-ChR2 (J), DR-NAC-Arch (K), and DR-NAC-ChR2 (L) subjects and their respective controls. Choice difficulty negatively modulates green-light effects in the DR-Arch and DR-NAC-Arch groups. Choice difficulty positively modulates blue-light effects in the DR-ChR2 and DR-NAC-ChR2 groups (10 mice; Wilcoxon signed-rank test; ∗p < 0.05; ∗∗p < 0.01).
Data are represented as mean ± SEM.
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